Epigenetics can be described as a stable alteration in gene expression potential that takes place during development and cell proliferation, mediated by mechanisms other than alterations in the primary nucleotide sequence of a gene. Three related mechanisms that cause epigenetic alteration in gene expression are DNA methylation, histone code changes and RNA interference.
DNA methylation is the main epigenetic modification in humans. It is a chemical modification of DNA performed by enzymes called methyltransferases, in which a methyl group (m) is added to specific cytosine (C) residues in DNA. In mammals, methylation occurs only at cytosine residues adjacent to a guanosine residue, i.e. at the sequence CG or at the CpG dinucleotide. In normal cells, methylation occurs predominantly in regions of DNA that have few CG base repeats, while CpG islands, regions of DNA that have long repeats of CG bases, remain non-methylated. Gene promoter regions that control protein expression are often CpG island-rich. Aberrant methylation of these normally non-methylated CpG islands in the promoter region causes transcriptional inactivation or silencing of certain functional genes in human cancers (Jones 2002).
Diagnostic markers for cancer detection have been described. One can distinguish between immunological markers and genetic markers. Genetic markers are based on detection of mutation in distinct genes, in particular in tumor suppressor genes. More recently, DNA methylation markers have been evaluated as potential genetic markers for detection of cancer because they offer certain advantages when compared to mutation markers. One of the most important features is that they occur at the early stages of cancer development and in many cases are tissue- and tumor-type specific (Esteller et al. 2001). A further advantage is that the methylation profile is preserved in purified isolated DNA and methylation changes appear to precede apparent malignancy in many cases. In addition, methylation markers may serve predictive purposes as they often reflect the sensitivity to therapy or duration of patient survival. All of these features find their application in improved cancer detection and therapy.
An early diagnosis is critical for the successful treatment of many types of cancer. The traditional methods of diagnosis (such as cytology, histopathology, immunohistochemistry, serology, and so on) are useful, but molecular markers can further subclassify the tumors and identify predisposition to cancer. If the exact methylation profiles of tumors are available and drugs targeting the specific genes are obtainable, then the treatment of cancer could be more focused and rational. Therefore, the detection and mapping of novel methylation markers is an essential step towards improvement of cancer prevention, screening and treatment.
Each year in the U.S. and EU, bladder cancer is diagnosed in >160,000 men and results in >48,000 deaths. While the five-year survival rate for early-stage bladder cancer is high, over 25% present with advanced disease and around 70% experience recurrence or progression following treatment. Urine cytology and cystoscopy are the current standard-of-care for bladder cancer detection and surveillance. Cystoscopy is highly sensitive but is invasive, expensive and causes significant patient discomfort. Urinary cytology is the most widely used method for non-invasive detection with up to 100% specificity. Unfortunately, this method is limited by its sensitivity, which is especially poor for low-grade bladder tumours.
Several methods have been reported for the detection of tumour cells in voided urine. However, none of these urinary tests can replace cystoscopy due to their poor specificity. Combining different methods of bladder cancer detection has been shown to improve sensitivity but unfortunately at the expense of specificity (Lotan Y et al., 2003).
Activating mutations in the fibroblast growth factor receptor 3 (FGFR3) gene have been reported in >50% of primary bladder tumors (van Rhijn B W G et al., 2003). Most of the somatic mutations found in bladder cancer are identical to germ line mutations responsible for skeletal disorders such as thanatophoric dysplasia and achondroplasia (van Rhijn B W G et al., 2002). It has been reported that FGFR3 mutations are very frequent in bladder tumors of low stage and grade, indicating that they occur much more frequently in superficial bladder cancer than in invasive bladder cancer (Billerey C et al., 2001). Recently the development of a new method for FGFR3 mutation analysis based on the detection of single nucleotide changes has been described by van Oers et al. With this method, the nine most common mutations can be detected in one assay simultaneously.